Effects of in vitro growth phase on the pathogenesis of Salmonella typhimurium in mice.

The growth phase of a bacterial (Salmonella typhimurium) culture was shown to have pronounced effects on the pathogenic properties of the harvested bacteria. Salmonellae obtained from a culture in primary (exponential) growth phase (PP) were more readily cleared from the blood and more readily killed by phagocytes than were salmonellae obtained from a more slowly growing secondary growth phase (SP) culture. PP salmonellae were observed to cause death of mice sooner than SP salmonellae. This appeared to be because the more rapid growth of PP, as compared to SP, salmonellae continued in the liver and spleen for several hours following intravenous injection, and more than compensated for their high in vivo death rate. As a result, within 4 h there were approximately 10-fold more live salmonellae in the spleens and livers of mice that had received PP, as compared to SP, salmonellae. This 10-fold difference was maintained until the death of the mice, indicating that after the first 4 h post-inoculation, the net in vivo growth of the salmonellae was the same regardless of their growth phase in the inoculating culture. This transition between PP and SP salmonellae occurred long before a dense stationary phase culture was obtained. Salmonellae grown in minimal media exhibited the biological properties of SP salmonellae and never entered as rapid a growth phase as did salmonellae in complete media.

Genetic mapping of novel virulence determinants of Salmonella typhimurium to the region between trpD and supD.

Salmonella typhimurium strains are known to vary greatly in their virulence for mice. A few of the genes affecting their virulence have been described. In this report we have localized at least two genes that affect the ability of S. typhimurium to grow in BALB/cByJ to a 6 unit section of the salmonella chromosome which does not contain any previously described virulence determinants. The genetic mapping was done by interrupted matings using Hfr strains made in a virulent LT2 strain. The Hfr strains were constructed by inserted the plasmid F'(TS)114 lac+ Tn::10 into the LT2 chromosome at specific sites through homologous recombination with chromosomal Tn10s. Short interrupted matings to an avirulent LT2 strain in either direction through the portion of the chromosome from trpD at 34 units to supD at 40 units resulted in transconjugants which were fully virulent. Since we also found several transconjugants with intermediate virulence it appears that more than one virulence gene may exist in this area of the chromosome. The mechanisms of action of these genes are not known.

Salmonella typhimurium virulence genes necessary to exploit the Itys/s genotype of the mouse.

The studies described in this paper provide evidence that the pathogenesis of Salmonella typhimurium in mice is dependent on interactions between particular genotypes of the infected mice and the infecting Salmonella strain. This conclusion was based on data obtained by infecting a panel of BXD recombinant inbred mice with each of three S. typhimurium strains. One of the S. typhimurium strains was a transconjugant (WB500) produced in an interrupted mating between the two other strains, one (SR-11) of high and the other (LT2-Z) of low virulence for BALB/c mice. We found that strain WB500 appeared to have inherited from SR-11 a gene (or genes) which was required to exploit the Itys/s genotype in mice. However, WB500 apparently lacked other SR-11 virulence gene(s), whose effect on the in vivo growth of SR-11 was independent of the Ity genotype of the mouse.

The ability of Salmonella typhimurium to produce the siderophore enterobactin is not a virulence factor in mouse typhoid.

One of the nonspecific defense mechanisms of higher animals is their ability to limit iron availability to infecting bacteria. Thus it has been argued that all pathogenic bacteria must have special mechanisms to obtain iron in the host environment. Salmonella typhimurium is known to produce a siderophore, enterobactin, with which it can obtain iron from host transferrin. Previous studies have indicated that the production of this molecule is necessary for the ability of intraperitoneally injected. S. typhimurium cells to cause mouse typhoid, a largely intracellular infection. We have reexamined this finding with wild-type S. typhimurium and isogenic strains carrying the nonenterobactin-producing mutation ent-1 or ent-7. Our findings demonstrate that, although enterobactin production is necessary for growth in normal mouse serum, it does not affect the ability of S. typhimurium to cause mouse typhoid. Based on these findings and published results of other investigators on the role of siderophores in intracellular pathogens, a more comprehensive investigation of the importance of siderophores in intracellular infections may be warranted.